1. Field of the Invention
The present invention is directed to a diagnostic test method and device for determining the presence of antigenic or enzmatic materials in body fluids. The instant invention relates broadly to a novel color indicating device and test method for immunological analysis of body materials and more particularly to pregnancy determination.
2. Description of the Prior Art:
A variety of physiological conditions or states, both normal and pathological, have been clinically diagnosed by a number of techniques, the more prominent being categorized as chemical, biological assay, or immunological. In many situations, the condition which is the object of the medical practitioner's determination is distinguished by the presence of one or more compounds, which because of the complexity or number of moieties present, obviates the use of chemical methods of analysis. In such instances biological assay methods, employing test animals, or immunological test procedures have been developed. It is the latter category of diagnostic means with which the present invention is concerned.
Analytical immunological or immunochemical test procedures are generally based upon an antigen-antibody reaction to determine the presence or absence of an antigen, antibody or enzyme in a body fluid ("Body fluid", as used herein, includes blood, serum, plasma, urine, cerebro-spinal fluid and saliva).
Pregnancy diagnostic tests have used both biological assay methods employing laboratory test animals and immunological test procedures. The former require the use of specific animals such as rabbits in the Friedman test or mice in the Aschheim-Zondek test. Such tests suffer from a number of disadvantages, such as, maintenance of large numbers of animals and attending facilities, particular skills in performing and analyzing the results of the tests, and the several day period required to obtain results of such tests, to name only a few.
In recent years, such biological tests have been supplanted by immunological or immunochemical tests offering many advantages. Namely, such tests may be performed in a physician's office, clinic or hospital, by technicians with less skill than that required with the biological tests. Results are usually obtained in shorter periods of time than with the older methods of diagnosis, and costs and other problems incurred in maintaining laboratory animals are avoided. In addition, immunological tests demonstrate greater specificity and sensitivity than bioassay techniques.
Most of the currently used immunological tests, particularly those employed in pregnancy determinations, fall into two broad categories, inhibition tests and direct agglutination tests. Both of these techniques, when applied to pregnancy tests, detect human chorionic gonadotropin (HCG) in the urine of pregnant women. Only in certain situations, representative of abnorml or pathological situations, is this hormonal substance encountered in non-pregnant women. The inhibition test employs erythrocyte cells or latex particles as substrate or carrier particles which are coated with HCG. A specimen of the urine being tested is initially incubated with anti-HCG. If the urine contains HCG bind to the anti-HCG present and inhibit any further binding of the antibody. An aliquot of the HCG-coated substrate particles is then added. If agglutination occurs, it indicates that the anti-HCG was free to passively agglutinate the substrate bound HCG and the conclusion is drawn that no HCG was present on the urine sample. If, however, no agglutination occurs, the inference may be drawn that HCG was present in the urine which effectively bound the anti-HCG, thus preventing interaction between the latter and the coated particles.
A direct agglutination test or latex agglutination test simply combines the specimen sample, suitably filtered and diluted with buffer solution, with a suspension of latex or other substrate particles coated with an antibody or antiserum fraction, such as anti-HCG. The presence of adequate HCG in the specimen sample will result in passive agglutination between the coated substrate particles and the HCG molecules.
Both types of tests may give inaccurate results due to false positives traceable to such things as impurities in the specimen sample and test reagents, excessive amounts of antibody coating, and too large or too small substrate particles to name but a few problems. One of the most frequent sources of error is related to the skills of the person interpreting the test. A lack of experience in interpreting test results will often lead to inaccurate results. Both categories of tests give end points which are somewhat equivocal, the latex agglutination test, for instance, indicating a positive test by a change from a milky white to a grainy white.
Many attempts have been made to develop more accurate, sensitive and definitive tests. U.S. Pat. No. 3,088,875 describes a method which incorporates a dye into a latex reagent in order to facilitate visualization of clumped or agglutinated particles. U.S. Pat. No. 3,236,732 describes a pregnancy test method in which a body fluid is combined with an antibody or antiserum of HCG and an indicator system which is broadly disclosed as an indicator material which in the presence of an antibody will agglutinate, precipitate, discolor, become colored or provide some other visible indication of the presence or absence of HCG. U.S. Pat. No. 3,862,302 discloses a single container, pregnancy diagnostic method employing stable pelletized reagents. U.S. Pat. No. 3,666,421 describes a diagnostic test slide which may be used advantageously to determine the presence of HCG in urine. The test card contains at least two dried reagent spots in close proximity which are intended to be reconstituted by addition of a liquid to be tested to form test reagents. One of the embodiments discloses the reagent spots being formed from HCG and an antiserum of HCG placed on a circle of contrasting color in order to aid in observing the test results.
Although certain advances have been made in the improvement in sensitivity, specificity, stability, and speed of immunological test procedures and the reagents and devices used therein, the techniques and materials developed heretofore have not provided a method and device which could be employed by one having little or no skill in performing such tests.